1. Field of the Invention
This invention relates generally to devices and methods used to test for and monitor the concentrations in solutions of sugars, amino acids and other compounds capable of complexing metal ions. More particularly, the present invention is directed to sensors which rely on metal coordination/chelation interactions between nucleophilic groups on targeted compounds and the release of protons, hydroxide ions or detectable ligands from metal ion complexes to provide detection and/or measurement of analyte compounds in aqueous, mixed aqueous-organic or organic solutions.
2. Description of Related Art
Many different devices and methods are presently being used to measure the concentration of various sugars and amino acids in a wide variety of solutions. Many industrial manufacturing and food processing systems require that the level of one or more sugars and/or amino acids be carefully monitored at various stages to insure desired quality of final products. For example, the varying glucose concentrations during fermentation processes are important process control parameters, and their continuous monitoring can improve the yield and quality of the fermentation product. In addition, there are a large number of situations where the amount of sugar in finished food stuffs and other sugar containing products must be determined. On line, continuous measurement is important to reduce the risk of contamination, labor costs and delays associated with off-line measurements. To be able to perform on-line measurements, there is a need for sterilizable sensors with rapid response times and high sensitivity, yet which also require minimum maintenance and calibration.
One of the most important uses for sugar analysis techniques is in the medical field where monitoring of sugar levels in biological fluids is critical to proper diagnosis and treatment of diabetes and other diseases. With respect to medical applications, glucose is by far the most important sugar, and diabetes is the most common disease for which glucose determinations are routinely conducted. Diabetes is a disease of the metabolic system that affects more than 14 million people in the United States and over 100 million people worldwide. It is characterized by an elevated blood-glucose concentration which is caused by a lack of the hormone insulin. Sugars are the primary source of metabolic energy, and the inability to self-regulate the levels of sugar metabolized by the body leads to many other medical problems, including but not limited to blindness, heart disease and kidney failure.
Treatment of diabetes involves monitoring of the patient's blood-glucose levels, with insulin injections being given when the glucose concentration rises above normal levels. A simple and accurate method for measuring blood-glucose concentrations is an essential cornerstone of any diabetes treatment protocol, since excessively high blood-glucose levels in diabetes patients can result in coma and even death. Frequent testing and insulin administration can significantly reduce long-term complications of diabetes. The vast majority of sensors which are used currently for glucose monitoring are based on enzymes such as glucose oxidase or glucose dehydrogenase. These enzyme-based sensors are simple to use and have relatively high sensing selectivity. They are widely used for one-time measurement of blood-glucose concentrations ex vivo. However, among the many drawbacks of enzyme-based sensors are that they are costly and have a short life time. The inherently unstable enzyme must be protected from extreme conditions during manufacturing and storage in order to preserve its catalytic activity. In addition, there have been a number of problems associated with the use of enzymes in implantable sensors used in systems for continuously monitoring blood-glucose levels in vivo, among them the fact that enzymes can elicit an immune response and are not stable to most sterilization methods.
The one-time or `spot` measurement of blood-glucose concentration ex vivo is also not optimal, as it requires collection of a blood sample, usually obtained by pricking the finger, which must be done at least several times each day. To avoid the need to subject the patient to this painful process for glucose monitoring, much effort has gone into identifying alternative sources for samples, such as subcutaneous tissue fluid, urine or saliva instead of blood, and for identifying less-painful and more efficient ways to obtain these samples, such as by transdermal extraction or using very thin needles. The development of new, highly sensitive and miniaturizable glucose monitoring technology will make some of these alternative methods more feasible for use at home by patients.
Continuous real time measurement of glucose concentrations is most desirable because it can be used for close monitoring and treatment. Studies indicate that medical outcomes are improved by more frequent, smaller insulin pulses. Continuous monitoring of glucose can also form part of a fully automatic insulin delivery system. A number of different sensor configurations have been proposed for use in either ex vivo or in vivo monitoring systems. Continuous glucose monitoring systems are described in a number of publications, for example by E. Wilkins and M. G. Wilkins (J. Biomed. Eng. 1983, Vol. 5, October, pp. 309-315) and J. Pickup (TIBTECH, July, 1993, Vol. 11, pp. 285-291).
Existing glucose sensing technologies exploit the ability of certain enzymes to selectively recognize glucose and catalyze a chemical reaction (Pickup, J. Trends in Biotechnology, 11, 285-291, (1993)). Many, for example, recruit glucose oxidase to catalyze the oxidation of glucose to gluconic acid and hydrogen peroxide, with electrochemical measurement of the latter. Potentiometric monitoring of gluconic acid production using a pH electrode or a field effect transistor (FET) is also possible. The enzyme-based sensors are simple to use and have relatively high sensing selectivity. They are widely used for one-time measurement of blood-glucose concentrations ex vivo. However, the enzyme system also has numerous disadvantages. These problems include high cost, difficulty in manufacturing, stability, both in ex vivo and in in vivo implantable devices (Alva et al. "Glucose-Oxidase Immobilized Electrode for Potentiometric Estimation of Glucose," Biosensors and Bioelectronics 6, 663-668, (1991); Shulga et al. "An Alternative Microbiosensor for Hydrogen-Peroxide Based on an Enzyme Field-Effect Transistor with a Fast-Response," Analytica Chimica ACTA 296, 163-170, (1994)). Also, the enzymes cause immunological responses and are difficult to sterilize for long term, continuous, real time measurement of glucose concentrations in vivo (Kerner et al. W., Kiwit, M., Linke, B., Keck, F. S., Zier, H. and Pfeiffer, E. F. "The Function of a Hydrogen Peroxide-Detecting Electroenzymatic Glucose Electrode is Markedly Impaired in Human Sub-cutaneous Tissue and Plasma," Biosensors and Bioelectronics 8, 473-482, (1993)). There is a need to develop sensitive and miniaturizable glucose monitoring devices which will make alternative methods of sample collection, such as from subcutaneous tissue fluid, more feasible for use at home by patients. The development of non-enzymatic approaches to glucose sensing is necessary in order to provide more effective management of diabetes, both for spot monitoring of glucose concentrations as well as for in vivo continuous monitoring.
A few nonenzymatic methods for measuring glucose have been proposed as alternatives to the above-described enzyme-based devices. U.S. Pat. No. 5,217,691 describes the use of boronic acids for the semiquantitative colorimetric determination of glucose. U.S. Pat. No. 4,371,374 discloses separating and quantitating glycosylated amino acids, peptides or mixtures thereof by treating a urine sample with a suitable boronic acid to complex the glycosylated compounds, separating them and analyzing the separated complexed material. U.S. Pat. No. 5,503,770 discloses a fluorescent boronic acid conjugate which emits fluorescence upon binding to saccharides. U.S. Pat. No. 5,244,562 discloses switching devices coated with a polymer imprinted with glucose and boronic acid.
Although some of the above-described sensors have shown promise, none have been found to be entirely satisfactory. In particular, the formation of covalent boronic acid-sugar complexes require several minutes to reach equilibrium, making their use in glucose sensors less attractive. There is a continuing need to develop robust sensor systems which can be used to accurately, simply and rapidly measure the amount of a particular sugar, amino acid or related compound which is present in solutions and other environments. The need is particularly apparent with respect to ex vivo and in vivo glucose determinations which are critical in medical diagnosis and treatment of various metabolic disorders, including diabetes. There is also a strong need to develop robust sensor technology for real-time monitoring of sugars, amino acids and other metabolites in manufacturing process environments.